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K Ogura

Publications and source records attributed to K Ogura.

At least 19 recordsLinked to original sources
PubMed1992

Peroxidase-amplified assay of sialidase activity toward gangliosides.

Sialidase assays were carried out with the substrate, ganglioside GD1a, coated onto enzyme immunoassay plate wells. Following the incubation of GD1a with sialidase from V. cholerae, the amount of ganglioside GM1 produced was measured as follows: cholera toxin B subunit conjugated to horseradish peroxidase was added to specifically bind to GM1, and then the amount of bound peroxidase was determined in a colorimetric enzymatic assay. In the absence of detergent, linearity for the detection of GM1 was 0 to 0.5 pmol per well, and the sensitivity for sialidase detection was about 3 fmol of product formed per minute. The addition of detergent (Triton CF-54) to the assay reduced the sensitivity and increased the amount of substrate required. Application of this assay for the detection of cell-derived neutral (pH 6.5) sialidase activities in the conditioned medium of human skin fibroblasts is described.

Chromatography, Gel↗
PubMed1992

Studies on geranylgeranyl diphosphate synthase from rat liver: specific inhibition by 3-azageranylgeranyl diphosphate.

Geranylgeranyl diphosphate synthase from rat liver was separated from farnesyl diphosphate synthase, the most abundant and widely occurring prenyltransferase, by DEAE-Toyopearl column chromatography. The enzyme catalyzed the formation of E,E,E-geranylgeranyl diphosphate (V) from isopentenyl diphosphate (II) and dimethylallyl diphosphate (I), geranyl diphosphate (III), or farnesyl diphosphate (IV) with relative velocities of 0.09:0.15:1. 3-Azageranylgeranyl diphosphate (VII), designed as a transition-state analog for the geranylgeranyl diphosphate synthase reaction, was synthesized and found to act as a specific inhibitor for this synthase, but not for farnesyl diphosphate synthase. Diphosphate V and its Z,E,E-isomer (VI) also inhibited geranylgeranyl diphosphate synthase, but the effect was not as striking as that of the aza analog VII. Specific inhibition of geranylgeranyl diphosphate synthase by VII was also observed in experiments with 100,000g supernatants of rat brain and liver homogenates which contained isopentenyl diphosphate isomerase and prenyltransferases including farnesyl diphosphate synthase as well as geranylgeranyl diphosphate synthase. For farnesyl:protein transferase from rat brain, however, the aza compound did not show a stronger inhibitory effect than E,E,E-geranylgeranyl diphosphate.

Animals↗
PubMed1992

Mitogenic effects of bacterial neuroaminidase and lactosylceramide on human cultured fibroblasts.

Exogenously added bacterial neuraminidase and lactosylceramide both stimulated the growth of cultured human skin fibroblasts. Neuraminidase (100 units/ml) increased DNA synthesis 1.9-fold and cell density 1.4-fold after 24 and 48 h, respectively, in culture. Treated fibroblasts contained less ganglioside NeuAc alpha 2-3Gal beta 1-4GlcCer (GM3), presumably due to neuraminidase-catalyzed hydrolysis to lactosylceramide. Addition of lactosylceramide (100 microM) to the fibroblast culture medium also increased DNA synthesis threefold within 24 h and cell density twofold after 48 h. These findings are compatible with a mechanism by which the proliferation of human fibroblasts is regulated by the relative levels of GM3 and lactosylceramide in the plasma membrane.

Antigens, CD↗
PubMed1992

Allelic loss of chromosome 17p in urothelial cancer: strong association with invasive phenotype.

Allelic loss of chromosome 17p with a mutated p53 gene on the remaining allele has been observed in various kinds of human cancers. To examine the significance of allelic loss of chromosome 17p in human urothelial cancer with special attention to the clinicopathological features, 49 tumors with various stages and grades from 43 cases (35 bladder cancers and 8 renal pelvic or ureteral cancers) were examined for loss of heterozygosity using 5 polymorphic probes on chromosome 17p. Thirty-seven cases were informative, and allelic loss of chromosome 17p was observed in 15 (41%) of them. In bladder cancers, the loss of 17p was observed with significantly higher frequency (p < 0.01) in cases with invasive (> or = pT2) tumors (7/10, 70%) than in cases with superficial (pTa or pT1) tumors (4/21, 19%). In renal pelvic or ureteral cancers, none of 2 superficial tumors and all of 4 invasive tumors showed the allelic loss. As to tumor grade, the allelic loss was observed in 1/9 (11%) for grade 1 cases, 6/18 (33%) for grade 2 cases, and 8/10 (80%) grade 3 cases (grade 1 versus 3, p < 0.01; grade 2 versus 3, p < 0.05). On the other hand, examination of clinical features, such as primary tumor site, tumor multiplicity or previous history of urothelial cancer did not significantly influence the frequency of the allelic loss. Our results suggest that the allelic loss of chromosome 17p is strongly associated with invasive phenotype in urothelial cancer. The results further indicate that the 17p deletion may represent a new genetic marker of malignant potentials in urothelial cancers.

Adult↗
PubMed1992

Farnesyl diphosphate synthase and solanesyl diphosphate synthase reactions of diphosphate-modified allylic analogs: the significance of the diphosphate linkage involved in the allylic substrates for prenyltransferase.

Diphosphate-modified substrates for prenyltransferase were synthesized and examined as substrates for the prenyltransferase reaction. They were dimethylallyl methylenediphosphonate, geranyl methylenediphosphonate, geranyl imidodiphosphate, geranyl phosphosulfate, farnesyl methylenediphosphonate, farnesyl imidodiphosphate, and farnesyl phosphosulfate. All of them except dimethylallyl methylenediphosphonate were accepted as substrates by solanesyl diphosphate synthase to give solanesyl diphosphate and the former four analogs were also accepted as substrates by farnesyl diphosphate synthase to give farnesyl diphosphate. The Km values of both enzymes for the methylenediphosphonate and imidodiphosphate analogs were comparable to those of the corresponding diphosphate substrates, but the phosphosulfate analogs showed much greater Km values than the diphosphate substrates. On the other hand, the Vmax values for these artificial substrates were all smaller than those for the corresponding natural substrates. Kinetic experiments with the analogs showed that the ionization-condensation-elimination mechanism proposed for the farnesyl diphosphate synthase reaction holds also for the solanesyl diphosphate synthase reaction and that the diphosphoryl structure, capable of chelating with divalent cations, is important topologically and kinetically rather than thermodynamically.

Alkyl and Aryl Transferases↗
PubMed1992

Chain length distribution of the products formed in solanesyl diphosphate synthase reaction.

Factors that affect the termination of isoprenoid chain elongation catalyzed by prenyltransferase were investigated. The chain-length distribution of reaction products of solanesyl diphosphate synthase [EC 2.5.1.11] homogeneously purified from Micrococcus luteus changed dramatically according to the concentration of the complex formed between isopentenyl diphosphate and Mg2+ (IPP-Mg) in the reaction mixture. However, the concentration of the complex between farnesyl diphosphate and Mg2+ (FPP-Mg), the priming substrate for this synthase, did not affect the product distribution, provided that the concentration of IPP-Mg was maintained at a certain level. Thus, the level of IPP-Mg is decisive in affecting the chain length distribution of the products of the prenyltransferase reaction, and the Mg(2+)-dependent variability of product specificity so far observed can now be understood in terms of the effect of IPP-Mg concentration.

Alkyl and Aryl Transferases↗
PubMed1992

Sensitization.

1. The transfusion effect, which had its greatest impact in 1985, disappeared in 1989. Since then, transfused and nontransfused groups have had identical 1-year graft survival rates. 2. The percentage of nontransfused recipients reached nearly 50% in 1992. The amount of pretransplant transfusion blood has dropped from 7 units in 1982 to below 5 units in 1992. Whole blood and packed red cells were the preferred products, comprising 98% of total transfusions in 1992. 3. The transfusion effect was observed in both sensitized and nonsensitized transfused recipients until 1988, although the effect was greater in the nonsensitized group. Since 1989, the sensitized group of transfused recipients had an even lower 1-year graft survival rate, while the nonsensitized group had a slightly better survival rate than those with no pretransplant transfusions. 4. Although washed red cells, frozen red cells, and frozen plasma constituted only a small percentage of total transfusions, any pretransplant transfusion of these blood products still produced a transfusion effect. Since 1982, patients who received washed red cells, frozen red cells, or frozen plasma had 8% higher 1-year graft survival rates since 1982 than those who had no transfusions, a difference still patient in 1990. 5. The percentages of broadly sensitized recipients decreased significantly in the last decade. Even recipients who had very specific antibodies (PRA 1-10%) had significantly lower graft survival rates than those with no antibodies (PRA = 0). Patients with PRA 1-10% and those with PRA 11-50% had almost identical graft survival curves and the same half-life, whether first or regrafted recipients. Only 0-PRA patients could be distinguished as "nonsensitized." 6. Once patients were sensitized, any time pretransplant, the current PRA values had less meaning for predicting graft survival than the peak (historical) PRA values. Those with peak PRA 51-100% had the same graft survival rates regardless of their current PRA values. Peak PRA was more important than current PRA for predicting graft status. 7. The deleterious effect of a positive flow cytometry crossmatch (FCXM) was obvious in both first and regrafted recipients. The positive group had about a 10% lower 1-year graft survival rate than the FCXM-negative group. 8. The significance of the positive FCXM outweighed the current PRA values, especially in regrafted recipients and first transplant recipients with peak PRA values. 9. FCXM seemed to work only on patients who had a chance of being sensitized by HLA stimulation. There was no positive FCXM effect in nontransfused recipients.(ABSTRACT TRUNCATED AT 400 WORDS)

B-Lymphocytes↗
PubMed1992

[Chromosomal survey of 1001 subfertile males: incidence and clinical features of males with chromosomal anomalies].

A chromosomal survey using the G-banding technique was performed on 1,001 subfertile males examined at Kyoto University Hospital between January, 1985 and April, 1991. Thirty-six of them had major chromosome anomalies (3.6%). The incidence of major anomalies for the 154 azoospermia patients, 326 oligozoospermia patients with a sperm density of less than 20 x 10(6)/ml, and 521 patients with normal sperm density was 10.4, 4.6, and 1.0%, respectively. The major anomalies found in azoospermic patients were associated with the sex chromosome, whereas autosomal anomalies were found in patients with oligozoospermia. The incidence of anomalies for patients with azoo- or oligozoospermia was significantly higher than that for patients with normal sperm density (p = 0.000003). All of the 12 patients with Klinefelter syndrome were azoospermic. However, the serum testosterone levels were within normal range in 10 of these patients. Testicular biopsies showed spermatogenic arrest in 5 of the 12 azoo- or oligozoospermic patients with structural chromosomal anomalies. Except for one patient, treatments such as varicocelectomy or medication were not effective in the patients with structural chromosomal anomalies. Chromosome studies are important in the evaluation of subfertile male patients with sperm densities less than 20 x 10(6)/ml. Disorders of the spermatogenic process may be directly related structural chromosomal anomalies in some of the patients.

Adult↗
PubMed1992

[Prophylactic effect of ulinastatin on cisplatin-induced renal disorders in lung cancer patients].

In order to determine whether ulinastatin (US) can prevent renal dysfunction due to cisplatin (CDDP) or not, we comparatively studied serum BUN, serum creatinine (Cr), creatinine clearance (Ccr), urine gamma-glutamyl transpeptidase (gamma-GTP) excretion, urine N-acetyl-beta-D-glucosaminidase (NAG) excretion, fractional excretion of Na (FENa) and urine beta 2-microglobulin excretion between one group of 10 patients receiving single treatment with CDDP and another group of 10 patients receiving combination treatment with CDDP+US, in 20 subjects with primary lung cancer. In the US combination treatment group, urine gamma-GTP excretion, urine NAG excretion, and FENa elevation were significantly inhibited. Urine beta 2-microglobulin excretion was inhibited. In summary, US was considered to have a preventive effect against renal toxicity due to CDDP.

Aged↗
PubMed1991

Molecular cloning and amino acid sequencing of rat liver class theta glutathione S-transferase Yrs-Yrs inactivating reactive sulfate esters of carcinogenic arylmethanols.

A cDNA containing the entire coding sequence for the subunit protein of rat liver class theta glutathione S-transferase (GST) Yrs-Yrs was isolated from a rat liver lambda gt11 cDNA library. The cDNA, designated GST theta-1, consisted of 1,258 bp which had an open reading frame of 732 bp encoding a polypeptide of 244 amino acid (AA) residues, including the leading AA Met to be removed on expression. The authenticity of the cDNA structure was supported by matching its deduced AA sequence with N-termini of Yrs and peptides obtained thereof by tryptic digestion as well as by CNBr cleavage. The deduced AA sequence of the subunit Yrs (M.W. 27,311) had only a weak homology (19-23%) with those of rat liver classes alpha, mu, and pi GST isozymes. Thus, the first evidence for the molecular cloning of the class theta GST was provided.

Amino Acid Sequence↗
PubMed1991

Purification of solanesyl-diphosphate synthase from Micrococcus luteus. A new class of prenyltransferase.

The activity of solanesyl-diphosphate synthase from Micrococcus luteus is stimulated by a high molecular mass fraction (HMF) which is separated from cell-free extracts of the same bacterium by DEAE-Toyopearl chromatography followed by Sephadex G-100 chromatography. By employing HMF in the assay procedure, solanesyl-diphosphate synthase was able to be purified to homogeneity and was found to be a homodimer with a monomeric molecular mass of 34 kDa. In contrast to hexaprenyl- and heptaprenyl-diphosphate synthases, which are composed of two easily dissociable components that are inactive unless combined, the homogeneously purified solanesyl-diphosphate synthase itself showed a catalytic activity, though weak, catalyzing the synthesis of both (all-E)-nonaprenyl-(solanesyl-) and (all-E)-octaprenyl diphosphate. HMF does not affect the stability of solanesyl-diphosphate synthase or Km values for isopentenyl diphosphate and farnesyl diphosphate, but it markedly increases Vmax values in a time-dependent manner. Several lines of evidence indicate that HMF contains a factor which binds to polyprenyl products and removes them out of the active site of enzyme to facilitate and maintain the turnover of catalysis.

Alkyl and Aryl Transferases↗
PubMed1991

Had-1, a uridine 5'-diphosphogalactose transport-defective mutant of mouse mammary tumor cell FM3A: composition of glycolipids, cell growth inhibition by lactosylceramide, and loss of tumorigenicity.

Glycolipid compositions of mouse mammary tumor cell FM3A and its Newcastle disease virus-resistant mutant cell, Had-1, which was also characterized as a defective mutant of UDP-galactose transport to Golgi apparatus, have been studied. The major neutral glycolipid in FM3A was Gal beta 1-4Glc beta 1-1Cer (LacCer) (95%) and the rest was Glc beta 1-1Cer. The concentration of neutral glycolipids in Had-1 was only about one-fifth of that in FM3A. GlcB1-1Cer in Had-1 accounted for 79% of neutral glycolipids and the rest was LacCer, the content of which was decreased to 4% of that in FM3A. Ganglioside patterns of the two cell lines were similar, although gangliosides with N-glycolylneuraminic acid were increased in Had-1 cells compared with that in FM3A cells. The presence of NeuAc alpha 2-3-Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc beta 1-1Cer, NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4GlcNAc beta 1-3Gal beta 1-4Glc beta 1-2Cer, GM3, and GD3 was demonstrated by thin-layer chromatography immunostaining. 125I-Labeled Newcastle disease virus bound only poorly to gangliosides extracted from either FM3A or Had-1 cells on a high performance thin-layer chromatography plate. The effects of glycolipids on the growth of the two cell lines were also studied. Had-1 cells were more sensitive to glycolipids added exogenously than FM3A cells. Addition of GM3 had a stimulative effect on cell growth of Had-1. LacCer, Gal beta 1-3GalNAc beta 1-4Gal beta 1-4Glc beta 4-1Cer, and Glc beta 1-1Cer inhibited the growth of Had-1 cells. LacCer was the most potent inhibitor. LacCer immobilized on the culture plate also inhibited the growth of Had-1 cells. The inhibitory effect was recovered completely overcome by transferring the cells to LacCer-free medium. Had-1 cells were not tumorigenic in C3H/He mice, and furthermore the tumorigenic activity of FM3A cells was suppressed by the prior administration of Had-1 cells.

Animals↗
PubMed1991

Formation of Z,E,E-geranylgeranyl diphosphate by rat liver microsomes.

Rat liver microsomes catalyzed the formation of A,E,E-geranylgeranyl diphosphate from farnesyl diphosphate and isopentenyl diphosphate in the presence of Triton X-100. Studies on product specificity using various primers such as Z,E-farnesyl diphosphate, E,E-farnesyl diphosphate, Z,E,E-geranylgeranyl diphosphate, E,E,E-geranylgeranyl diphosphate, Z,E,E,E-geranylfarnesyl diphosphate, and E,E,E,E-geranylfarnesyl diphosphate suggested that the microsomal dehydrodolichyl diphosphate synthase has such properties that it releases Z,E,E-geranylgeranyl diphosphate, the first intermediate, in the reactions with farnesyl diphosphate as the starting primer. Metabolic labeling of rat liver slices with [2-3H]mevalonic acid revealed the accumulation of E,E,E-geranylgeranyl (di)phosphates as well as dolichyl (di)phosphate (C85 and C90) and dehydrodolichol (C85 and C90), but no accumulation of Z,E,E-geranylgeranyl (di)phosphate or E,E-farnesyl (di)phosphate was detected. Microsomal enzyme preparations from mouse liver and hamster liver also produced Z,E,E-geranylgeranyl diphosphate from farnesyl diphosphate and isopentenyl diphosphate.

Alkyl and Aryl Transferases↗
PubMed1991

Variable product specificity of microsomal dehydrodolichyl diphosphate synthase from rat liver.

Several detergents activated microsomal dehydrodolichyl diphosphate synthase of rat liver, but the chain length of products shifted downward from C90 and C95 with increasing concentration of the detergents. Maximum activation was observed at the concentration of 2% Triton X-100, 30 mM octyl glucoside, 30 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, and 10 mM deoxycholate with the product chain length being C80-C85, C65-C75, C70-C75, and C55-C65, respectively. The activity of Triton X-100 solubilized enzyme was decreased by asolectin, phosphatidylethanolamine, and phosphatidylcholine. The chain lengths of products formed in the presence of these phospholipids were C85 and C90. In the presence of both phosphatidylcholine and Mg2+ the solubilized enzyme was able to produce C90 and C95 dehydrodolichyl diphosphates like native microsomal enzyme. Microsomal enzyme preparations from rat liver, brain, and testis catalyzed the formation of dehydrodolichyl diphosphates with the same chain lengths as those of the natural dolichols occurring in individual tissues. The chain length distribution of dehydrodolichyl products by (rat liver) microsomes also depended on the concentration of substrates. Not only did increasing the concentration of isopentenyl diphosphate lead to longer chain product, but decreasing that of farnesyl diphosphate increased product chain length.

Alkyl and Aryl Transferases↗
PubMed1991

Uptake and metabolism of radiolabelled GM1-ganglioside in skin fibroblasts from controls and patients with GM1-gangliosidosis.

The uptake and metabolism of [3-3H-sphingosine]GM1-ganglioside was measured in cultured skin fibroblasts from controls and patients with infantile, juvenile and adult GM1-gangliosidosis. When dissolved in medium with phosphatidylserine, GM1-ganglioside was efficiently taken up by cultured skin fibroblasts and transferred into lysosomes. A linear increase in GM1-ganglioside endocytosis was shown with phosphatidylserine concentrations of up to 40 micrograms/ml. A pulse-chase study revealed that [3H]GM1-ganglioside was metabolized to GM2-ganglioside, GM3-ganglioside, ceramide dihexoside, ceramide monohexoside, ceramide and sphingosine. Sphingosine was recycled to sphingomyelin. In a 20-h pulse study, cell lines from patients with GM1-gangliosidosis of infantile, juvenile and adult types hydrolysed 2-5%, 20-44% and 54-58% of the total endocytosed GM1-ganglioside respectively. These values were lower than in control cells (64.17 +/- 5.43% (n = 10]. The hydrolysis rates of exogenous [3H]GM1-ganglioside in cultured fibroblasts from patients with various types of GM1-gangliosidosis closely reflected the clinical severity.

Cells, Cultured↗
PubMed1991

Differential effects of pentobarbital on intracerebral arterioles and venules of rats in vitro.

The vasoactive properties of pentobarbital (PB) were studied in intracerebral arterioles and venules (diameter, 30-90 microns). These vessels mediate changes in cerebrovascular resistance and capacitance, respectively. Mean control vessel diameters of arterioles and venules at pH 7.3 were 53.9 +/- 2.8 microns and 78.4 +/- 4.3 microns, respectively. Both arterioles and venules dilated when the pH of the extraluminal solution was lowered to 6.8 and constricted when the pH was raised to 7.6. PB, 10(-6) to 10(-2) mol/L, dilated intracerebral arterioles in a dose-dependent manner at pH 7.3, reaching a maximal dilation of 129.7 +/- 3.1% of control diameter at a dose of 10(-3) mol/L. In contrast, PB at 10(-6) to 10(-2) mol/L failed to produce significant changes in the diameter of intracerebral venules. In addition, PB at 10(-3) mol/L significantly inhibited arteriolar constriction induced by KCl (120 mmol/L), but not venular constriction. The present study suggests that intracerebral venules are relatively less responsive to PB than cerebral arterioles and peripheral veins. In addition to its effect on cerebral metabolism. PB may act to redistribute venous blood volume from cerebral veins to more responsive peripheral veins, thereby decreasing intracranial blood volume and intracranial pressure.

Animals↗